The goal of the MLPCN is to integrate high-throughput chemical approaches with state-of-the art genetics, (cellular, molecular and in vivo biology in a multi-disciplinary effort to discover of proof-of-concept (POC) molecular probes for cell and in vivo systems. Probes help transform biomedical advances to impact on public health and quality of life. Production capacity at The Scripps Center in the pilot MLSCN phase was demonstrated by publishing >68 assays in PUBCHEM, optimizing POC probes for 5 molecular targets and peer-reviewed publication of probes with low nanomolar potency, selectivity, and in vivo efficacy, to define novel biological and therapeutic targets. The Center sustained full production capabilities for assay development and for ?HTS implementation in 1536-well format (>20/year). >80 data sets were published in PUBCHEM at a rate of 2 full library assays/month. In the past 12 months alone, 27 ?HTS campaigns including 20 primary cell-based screens) for 18 molecular targets in 8 different target classes, using 8 detection formats (and from 13 external PIs) published. Internal discovery projects were screened on a single compound library of >600,000, exceeding MLPCN requirements. These data were achieved in well volumes (of 5-10 ?l at low cost. The Center spares NIH compound collection and controls cost using 2 ?l of compound (solution per year for 24 primary screens, and requiring only 5 ?l for hit-picks, reconfirmation and 10 point titrations. Formats include ion flux and GPCR assays, reporter gene, transcription factor and nuclear receptor assays, enzyme assays, protein-protein and protein-RNA interactions and phenotypic screens. The Center currently supports hit-to-probe chemical synthesis across 8 projects/year by resynthesis, chemi-informatics, medicinal chemistry, secondary and specificity assay implementation and screening pharmacokinetics/ metabolism studies for efficient probe optimization. The Center developed novel technologies for high throughput selectivity profiling across gene families and enzymes for optimizing target-selective probes. This Comprehensive Center supports production discovery of POC probes for most target classes by meeting 3 (Aims. 1: Develop 25 assays/year to HTS readiness. 2: Implement 25 ?HTS screens/year at 300-500,000 individual compounds and publish quality-assured data to PUBCHEM. 3: Optimize screening hits to probes or POC in cells and/or in vivo for 10-15 programs/year. The Center provides public data, tools and resources hat enhance the success of NIH Roadmap and impact on human health.